Problem Area: Severe pulmonary hypertension (PH) is a rare but deadly disease. This orphan disease is characterized by progressive increase in pressure in pulmonary circulation and vascular remodeling of small size pulmonary arteries that include proliferation of endothelial cells (ECs). Currently, there are few therapeutic options, and those available have poor efficacy. Background: Previous animal experiments and some observational studies in humans suggest that estradiol (E2) may attenuate the development of PH; however, the animal models employed do not mimic human PH, the clinical findings are ambiguous, and whether the effects of E2 are mediated by E2 per se or via its downstream metabolites has not been examined. Also, it is unclear whether direct or indirect (via metabolites) effects of E2 on pulmonary EC growth mediate the putative beneficial effects of estradiol in PH. Preliminary Studies: Estradiol is metabolized by CYP1A1 to 2-hydroxyestradiol (2HE) and 2HE is converted to 2-methoxyestradiol (2ME) via COMT. In a number of studies we have shown that 2ME inhibits vascular (media) remodeling in vivo and attenuates the development of monocrotaline-induced PH. Estradiol and 2ME have divergent effects on ECs; however, their effects on endothelial remodeling in severe PH have not been studied. Hypothesis: We hypothesize that estradiol attenuates severe PH via a mechanism that involves its metabolism to 2- methoxyestradiol and that E2 and 2ME differently affect EC proliferation and endothelial remodeling. Thereby, 2ME may offer highly efficacious protection in severe PH. To test our hypothesis we will use a rat model of severe PH that mimics the key alterations seen in the severe form of PH in humans. Specific Aims: We will address the following questions: 1) Does endogenous/exogenous estradiol attenuate the development of severe PH by a mechanism that involves its downstream metabolites, and does 2ME attenuate the development of severe PH? 2) Do alterations of E2 metabolism affect EC proliferation and do E2 and 2ME differently affect the proliferation of ECs from control and severe PH animals? Significance: This project will provide information that may ultimately lead to the development of safe and effective modalities for the treatment of severe pulmonary hypertension in humans. [unreadable] [unreadable]